Lymphocyte development is a dynamic yet tightly regulated process to give rise to T- and B- lymphocytes, the major cellular components of our immune system. Genetic alterations in key regulatory genes and could and does often result in immune deficiency, autoimmunity, lymphoma, or leukemia. Our long term research goal is to understand molecular mechanisms underlying lymphocyte development and to provide novel strategies for diagnosis and treatment of lymphoid diseases. In this proposal, we will specifically address the function of HEB in T-cell development. HEB encodes a bHLH type of transcription factor which is highly (although not exclusively) expressed in T-lymphocytes. We have used gene targeting in mice to show that HEB plays an essential role in early stages of T cell development. Evidence indicates that this function of HEB requires collaboration with other regulatory molecules such as E2A, TCF, and CBF1alpha. To better understand the mechanism underlying HEB function and to gain insights into the regulatory pathways mediated by HEB, we propose the following experiments: 1) We will further define the function of HEB and its relationship with E2A and evaluate the potential role of HEB in the cell death pathway. These will be accomplished by using a retroviral-based cDNA vector in an adoptive transfer assay and/or conventional transgenic rescuing assay. 2) We will use ENU mutagenesis in mice to screen for modifiers of HEB. This forward genetic approach will lead to identification of regulatory molecules whose functions are linked with HEB. 3) We will characterize and map a novel mutations identified in an earlier screen and other mutations to be isolated from this screen. In addition, we will also initiate the positional and functional cloning process so that this research will eventually lead to isolating novel regulatory genes important for T-cell development. Although the proposed mutagenesis experiment represents a long term commitment, our immediate goal in the funding period is to identify and characterize several lymphoid specific mutations and place them into the HEB pathway. The ultimate goal is to isolate these genes based on their predetermined function in T-cell development.